Abstract Blood cancers account for approximately 10% of all malignancies. In addition, non-malignant bone marrow failure diseases such as aplastic anaemia, or immunodeficiency diseases such as severe combined immunodeficiency, are life-threatening diseases and allogeneic bone marrow transplantation (BMT) is a preferred curative therapy for these serious conditions. BMT outcomes are limited by transplant related complications, mainly graft-versus- host disease (GVHD) and opportunistic infections. Indeed, 15-20% of BMT patients will develop severe GVHD that is fatal, particularly when involving the GI tract. Current prevention and treatment of GVHD rely on the broad suppression of T cells and remains suboptimal. The initiation and maintenance of T cell responses are dependent on activities mediated by antigen-presenting cells (APC) but the types of APC that initiate GVHD and the factors that promote their function is currently unknown. This is the focus of this proposal. In particular, we will build on our preliminary data to test the hypothesis that lethal acute GVHD is initiated in the ileum. We will utilize cutting- edge mechanistic preclinical murine studies with parallel clinical analysis, using advanced flow cytometry, RNASeq, and REAP/CITE-seq to focus on antigen presentation in the gastrointestinal tract, identifying clinically tractable pathways that will prevent the development of lethal acute GVHD, whilst permitting GVL and pathogen- specific immunity that are initiated in primary and secondary lymphoid organs. The proposal will lead to new, rapidly testing therapeutic approaches to prevent GVHD based on the inhibition of disease initiation (i.e. alloantigen presentation) rather than broad T cell suppression in the late effector phase of disease, as is current practice.